Solvent selection for insoluble ligands, a challenge for biological assay development: A TNF-α/SPD304 study

Christos P. Papaneophytou, Anthi K. Mettou, Vagelis Rinotas, Eleni Douni, George A. Kontopidis

Research output: Contribution to journalArticlepeer-review

Abstract

Many active compounds may be excluded from biological assays due to their low aqueous solubility. In this study, a simple method for the determination of the solubility of compounds containing aromatic rings is proposed. In addition to DMSO, five organic solvents for screening experiments of TNF-α inhibitors were explored. DMSO and PEG3350 were the most suitable for both protein stability and ligand-binding experiments. In addition, glycerol is a promising solvent for the screening of other compounds for which it might provide acceptable solubilization, due to its strong tendency to preserve the protein. Moreover, a fluorescence binding assay was developed using the TNF-α/SPD304 system, and a Kd of 5.36 ± 0.21 μM was determined. The results of this study could be used for the future screening of potential TNF-α inhibitors, while the protocols developed in this work could be applied to other proteins.

Original languageEnglish
Pages (from-to)137-141
Number of pages5
JournalACS Medicinal Chemistry Letters
Volume4
Issue number1
DOIs
Publication statusPublished - 10 Jan 2013

Keywords

  • aqueous solubility
  • fluorescence binding assay
  • insoluble ligands
  • protein stability
  • solvent selection

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